Cyanide-induced neurotoxicity is accompanied by destabilization of neuronal Ca+ homeostasis, triggering an uncontrolled, sustained rise in cytosolic free Ca2+ which in turn activates several neurotoxic pathways. This study will characterize the Ca2+ dependent signal cascades activated by cyanide and determine the mechanism of the subsequent neuronal dysfunction and injury in two neuronal models, the PC12 cell and primary cultured cerebellar granule cells. This will include studying the effect of cyanide on the nitric oxide signal cascade, protein kinase C activation and translocation, and calmodulin/calmodulin-dependent protein kinase II. Cyanide's action on each Ca2+ dependent system will be correlated with alterations in neuronal biochemistry and injury. The mechanism underlying cyanide-induced excitotoxicity will also be studied. Direct interaction of cyanide with the NMDA receptor redox modulatory site will be related to the excitotoxic response. Additionally, the role oxidative stress (hydroperoxide generation), inositol (1,4,5) triphosphate cascade and potassium channels play in the cyanide-induced excitotoxic response will be addressed. Finally, the contribution of genomic activation to the neurotoxicity will be determined. This includes correlation of cyanide stimulated proto-oncogene (c-fos) expression with the cytotoxic response. Nerve growth factor differentiated PC12 cells will be used to study cyanide-induced apoptotic death and the influence of impaired Ca2+ regulation and sustained activation of intracellular signal transduction on the apoptotic process. The long-term goal of this project is to develop an in-depth understanding of the mechanism of toxicity of a xenobiotic in the CNS by relating sustained activation of Ca2+ dependent cascades to neuronal cytotoxic responses.